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Alekseeva AA, Kargov IS, Kleimenov SY, et al. Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max. Acta Naturae. 2015;7(3):55-64
Alekseeva, A. A., Kargov, I. S., Kleimenov, S. Y., Savin, S. S., & Tishkov, V. I. (2015). Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max. Acta naturae, 7(3), 55-64.
Alekseeva, A A, et al. "Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max." Acta naturae vol. 7,3 (2015): 55-64.
Alekseeva AA, Kargov IS, Kleimenov SY, Savin SS, Tishkov VI. Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max. Acta Naturae. 2015 Jul-Sep;7(3):55-64. PMID: 26483960; PMCID: PMC4610165.
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Acta Naturae. 2015 Jul-Sep;7(3):55-64.

Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max.

Acta naturae

A A Alekseeva, I S Kargov, S Yu Kleimenov, S S Savin, V I Tishkov

Affiliations

  1. A.N.Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninskiy Prospect, 33/2, Moscow,119071, Russia ; Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia.
  2. Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia ; Departament of Chemistry, M.V. Lomonosov Moscow State University, Leninskie Gory, 1/3, Moscow, 119991, Russia.
  3. A.N.Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninskiy Prospect, 33/2, Moscow,119071, Russia ; N.K. Koltsov Institute of Developmental Biology of the Russian Academy of Sciences, Vavilova St., 26, Moscow, 119334, Russia.
  4. A.N.Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninskiy Prospect, 33/2, Moscow,119071, Russia ; Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia ; Departament of Chemistry, M.V. Lomonosov Moscow State University, Leninskie Gory, 1/3, Moscow, 119991, Russia.

PMID: 26483960 PMCID: PMC4610165

Abstract

Recently, we demonstrated that the amino acid substitutions Ala267Met and Ala267Met/Ile272Val (Alekseeva et al., Biochemistry, 2012), Phe290Asp, Phe290Asn and Phe290Ser (Alekseeva et al., Prot. Eng. Des. Select, 2012) in recombinant formate dehydrogenase from soya Glycine max (SoyFDH) lead to a significant (up to 30-100 times) increase in the thermal stability of the enzyme. The substitutions Phe290Asp, Phe290Asn and Phe290Ser were introduced into double mutant SoyFDH Ala267Met/Ile272Val by site-directed mutagenesis. Combinations of three substitutions did not lead to a noticeable change in the catalytic properties of the mutant enzymes. The stability of the resultant triple mutants was studied through thermal inactivation kinetics and differential scanning calorimetry. The thermal stability of the new mutant SoyFDHs was shown to be much higher than that of their precursors. The stability of the best mutant SoyFDH Ala267Met/Ile272Val/Phe290Asp turned out to be comparable to that of the most stable wild-type formate dehydrogenases from other sources. The results obtained with both methods indicate a great synergistic contribution of individual amino acid substitutions to the common stabilization effect.

Keywords: Glycine max; formate dehydrogenase; multi-point mutants; protein engineering; rational design; stability; stabilization; synergistic effect

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